Abstract

Aims Fourier transform infrared (FT-IR) imaging is increasingly being applied to biomedical specimens, but strong IR absorption
by water complicates live cell imaging. This study investigates the viability of adherent epithelial cells maintained for
short periods under mineral oils in order to facilitate live cell spectroscopy using FT-IR with subsequent imaging.

Methods The MGH-U1 urothelial or CaCo2 colorectal cancer cell lines were grown on plastic surfaces or mid-range infrared transparent
windows. Medium in established cultures was replaced with paraffin mineral oil, or Fluorolube, for up to 2 h, and viability
assessed by supravital staining. Drug handling characteristics were also assessed. Imaging of preparations was attempted by
reflectance and transmission using a Varian FT-IR microscope.

Results Cells covered by mineral oil remained viable for 2 h, with recovery into normal medium possible. MTT ((3-(4,5-dimethylthlazol-2-yl)-2,5-diphenyl
tetrazolium) conversion to crystalline formazan and differential patterns of drug uptake were maintained. The combination
of a calcium fluoride substrate, Fluorolube oil, and transmission optics proved best for spectroscopy. Spectral features were
used to obtain images of live cells.

Conclusions The viability of cells overlaid with IR transparent oils was assessed as part of a technique to optimise conditions for FT-IR
imaging. Images of untreated cells were obtained using both reflectance and transmission. This represents an effective means
of imaging live cells by IR spectroscopy, and also means that imaging is not necessarily a terminal event. It also increases
options for producing images based on real-time biochemistry in a range of in vitro experimental and ‘optical biopsy’ contexts.